A SIMULTANEOUS ASSAY OF THE DIFFERENTIATING AGENTS, PHENYLACETIC ACIDAND PHENYLBUTYRIC ACID, AND ONE OF THEIR METABOLITES, PHENYLACETYL-GLUTAMINE, BY REVERSED-PHASE, HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

This paper describes a reversed-phase, high performance liquid chromat ographic (HPLC) method for the isolation, detection and quantification of phenylacetic acid, phenylbutyric acid and one of their metabolites , phenylacetylglutamine, from serum. The compounds are initially extra cted from serum after protein precipitation with perchloric acid. The solution is neutralized with potassium bicarbonate and injected onto a C-18 column. The compounds are then differentially eluted using an in creasing gradient of acetonitrile. Ionization of the two acids is supp ressed by adding phosphoric acid to the mobile phase. The compounds ar e detected by UV absorbance at 208 nm. The assay yields a lower limit of detection of 2 mu g/ml and is linear to concentrations as high as 2 ,000 mu g/ml. Between 20 and 1,000 mu g/ml the interassay percent coef ficient of variation is less than 10%.